{
"METABOLOMICS WORKBENCH":{"STUDY_ID":"ST002488","ANALYSIS_ID":"AN004062","VERSION":"1","CREATED_ON":"February 21, 2023, 11:50 am"},

"PROJECT":{"PROJECT_TITLE":"Simultaneous targeting of PD-1 and IL2Rβγ with radiation therapy to inhibit pancreatic cancer growth and metastasis","PROJECT_SUMMARY":"In pancreatic ductal adenocarcinoma (PDAC) patients, we show that response to radiation therapy (RT) is characterized by increased IL2Rβ and IL2Rγ and decreased ILR2α expression. The bispecific aPD1-IL2v is a PD-1-targeted IL-2 variant (IL2v) immunocytokine with engineered IL-2 cis-targeted to PD-1 and abolished IL2Rα binding, which enhances tumor-antigen specific T cell activation while reducing regulatory T cell (Treg) suppression. Using aPD1-IL2v in orthotopic PDAC KPC-driven tumor models, we show marked improvement in local and metastatic survival along with profound increase in tumor-infiltrating polyfunctional CD8+ T cell subsets with a transcriptionally and metabolically active phenotype, and preferential activation of antigen-specific CD8+ T cells. In combination with single dose RT, aPD1-IL2v treatment results in a robust, durable expansion of polyfunctional CD8+ T cells, T cell stemness, tumor-specific memory immune response, natural killer (NK) cell activation, and decreased Tregs. These data show that aPD1-IL2v leads to profound local and distant response in PDAC.","INSTITUTE":"University of Colorado Denver","LABORATORY":"Lab of Angelo D'Alessandro in collaboration with lab of Sana Karam","LAST_NAME":"Haines","FIRST_NAME":"Julie","ADDRESS":"12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA","EMAIL":"julie.haines@cuanschutz.edu","PHONE":"3037243339"},

"STUDY":{"STUDY_TITLE":"Simultaneous targeting of PD-1 and IL2Rβγ with radiation therapy to inhibit pancreatic cancer growth and metastasis - mouse serum metabolomics","STUDY_SUMMARY":"C57BL/6 mice were orthotopically implanted with PK5L1940 cells. 8 Gy RT was administered 7 days post-implantation. PD1-IL2v and aCD25 dosed once per week beginning day 7 post-implantation. Spleens and lymph nodes were harvested 17 days post implantation and the sorted T-cells cultured in the presence of 13C6 glucose for 6h. This study contains longitudinal sampling of the cell culture supernatants.","INSTITUTE":"University of Colorado Denver","LAST_NAME":"Haines","FIRST_NAME":"Julie","ADDRESS":"12801 E 17th Ave, Room 1303, Aurora, Colorado, 80045, USA","EMAIL":"julie.haines@cuanschutz.edu","PHONE":"3037243339"},

"SUBJECT":{"SUBJECT_TYPE":"Mammal","SUBJECT_SPECIES":"Mus musculus"},
"SUBJECT_SAMPLE_FACTORS":[
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-1",
"Factors":{"Factor":"Untrx","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-1_r16-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-2",
"Factors":{"Factor":"Untrx","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-2_r23-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-3",
"Factors":{"Factor":"Untrx","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-3_r48-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-4",
"Factors":{"Factor":"RT","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-4_r49-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-5",
"Factors":{"Factor":"RT","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-5_r28-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-6",
"Factors":{"Factor":"RT","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-6_r4-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-7",
"Factors":{"Factor":"IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-7_r53-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-8",
"Factors":{"Factor":"IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-8_r20-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-9",
"Factors":{"Factor":"IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-9_r35-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-10",
"Factors":{"Factor":"RT+IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-10_r41-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-11",
"Factors":{"Factor":"RT+IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-11_r46-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-12",
"Factors":{"Factor":"RT+IL2v","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-12_r36-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-13",
"Factors":{"Factor":"aCD25","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-13_r14-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-14",
"Factors":{"Factor":"aCD25","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-14_r10-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-15",
"Factors":{"Factor":"aCD25","Timepoint":"5 min"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-15_r40-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-16",
"Factors":{"Factor":"Untrx","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-16_r27-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-17",
"Factors":{"Factor":"Untrx","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-17_r31-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-18",
"Factors":{"Factor":"Untrx","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-18_r38-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-19",
"Factors":{"Factor":"RT","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-19_r54-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-20",
"Factors":{"Factor":"RT","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-20_r39-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-21",
"Factors":{"Factor":"RT","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-21_r19-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-22",
"Factors":{"Factor":"IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-22_r1-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-23",
"Factors":{"Factor":"IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-23_r18-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-24",
"Factors":{"Factor":"IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-24_r58-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-25",
"Factors":{"Factor":"RT+IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-25_r60-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-26",
"Factors":{"Factor":"RT+IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-26_r55-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-27",
"Factors":{"Factor":"RT+IL2v","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-27_r2-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-28",
"Factors":{"Factor":"aCD25","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-28_r17-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-29",
"Factors":{"Factor":"aCD25","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-29_r22-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-30",
"Factors":{"Factor":"aCD25","Timepoint":"1 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-30_r6-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-31",
"Factors":{"Factor":"Untrx","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-31_r37-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-32",
"Factors":{"Factor":"Untrx","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-32_r3-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-33",
"Factors":{"Factor":"Untrx","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-33_r24-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-34",
"Factors":{"Factor":"RT","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-34_r44-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-35",
"Factors":{"Factor":"RT","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-35_r42-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-36",
"Factors":{"Factor":"RT","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-36_r26-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-37",
"Factors":{"Factor":"IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-37_r50-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-38",
"Factors":{"Factor":"IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-38_r13-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-39",
"Factors":{"Factor":"IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-39_r12-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-40",
"Factors":{"Factor":"RT+IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-40_r29-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-41",
"Factors":{"Factor":"RT+IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-41_r51-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-42",
"Factors":{"Factor":"RT+IL2v","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-42_r59-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-43",
"Factors":{"Factor":"aCD25","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-43_r52-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-44",
"Factors":{"Factor":"aCD25","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-44_r7-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-45",
"Factors":{"Factor":"aCD25","Timepoint":"2 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-45_r45-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-46",
"Factors":{"Factor":"Untrx","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-46_r43-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-47",
"Factors":{"Factor":"Untrx","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-47_r57-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-48",
"Factors":{"Factor":"Untrx","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-48_r25-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-49",
"Factors":{"Factor":"RT","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-49_r5-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-50",
"Factors":{"Factor":"RT","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-50_r8-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-51",
"Factors":{"Factor":"RT","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-51_r34-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-52",
"Factors":{"Factor":"IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-52_r47-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-53",
"Factors":{"Factor":"IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-53_r30-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-54",
"Factors":{"Factor":"IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-54_r56-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-55",
"Factors":{"Factor":"RT+IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-55_r15-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-56",
"Factors":{"Factor":"RT+IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-56_r9-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-57",
"Factors":{"Factor":"RT+IL2v","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-57_r33-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-58",
"Factors":{"Factor":"aCD25","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-58_r11-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-59",
"Factors":{"Factor":"aCD25","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-59_r21-"}
},
{
"Subject ID":"-",
"Sample ID":"AG-23-TS-60",
"Factors":{"Factor":"aCD25","Timepoint":"6 hr"},
"Additional sample data":{"RAW_FILE_NAME":"AG-23-TS-60_r32-"}
}
],
"COLLECTION":{"COLLECTION_SUMMARY":"C57BL/6 mice were orthotopically implanted with PK5L1940 cells. 8 Gy RT was administered 7 days post-implantation. PD1-IL2v and aCD25 dosed once per week beginning day 7 post-implantation. Spleens and lymph nodes were harvested 17 days post implantation and homogenized to a single cell suspension by mashing through a 70 uM cell strainer. For spleen processing, the erythrocytes were lysed with ACK (ammonium-chloride-potassium) lysis buffer for 3 min at RT. CD8 T cells were sorted with a CD8-negative selection Stemcell isolation kit following manufacturer instructions.","SAMPLE_TYPE":"Cell culture supernatant"},

"TREATMENT":{"TREATMENT_SUMMARY":"CD8+ T cells were cultured for 6h in the presence of 25 mM U-13C-glucose in glucose-free RPMI. Media aliquots were taken at 5 min, 1h, 2h, 6h and snap frozen."},

"SAMPLEPREP":{"SAMPLEPREP_SUMMARY":"Supernatant aliquots were thawed on ice then a 20 uL aliquot treated with 480 uL of cold 5:3:2 MeOH:acetonitrile:water. Samples were vortexed 30 min at 4 degrees C then supernatants clarified by centrifugation (10 min, 10,000 g, 4 degrees C) and transferred to autosampler vials.","PROCESSING_STORAGE_CONDITIONS":"4℃","EXTRACT_STORAGE":"-80℃"},

"CHROMATOGRAPHY":{"CHROMATOGRAPHY_SUMMARY":"Negative C18","CHROMATOGRAPHY_TYPE":"Reversed phase","INSTRUMENT_NAME":"Thermo Vanquish","COLUMN_NAME":"Phenomenex Kinetex C18 2.1 x 150 mm, 1.7 um","SOLVENT_A":"95% water 5% acetonitrile 1 mM ammonium acetate","SOLVENT_B":"95% acetonitrile 5% water 1 mM ammonium acetate","FLOW_GRADIENT":"0-0.5 min 0% B, 0.5-1.1 min 0-100% B, 1.1-2.75 min hold at 100% B, 2.75-3 min 100-0% B, 3-5 min hold at 0% B","FLOW_RATE":"450 uL/min","COLUMN_TEMPERATURE":"45","SAMPLE_INJECTION":"10 uL"},

"ANALYSIS":{"ANALYSIS_TYPE":"MS"},

"MS":{"INSTRUMENT_NAME":"Thermo Q Exactive Orbitrap","INSTRUMENT_TYPE":"Orbitrap","MS_TYPE":"ESI","ION_MODE":"NEGATIVE","MS_COMMENTS":"Resolution 70,000, scan range 65-900 m/z, maximum injection time 200 ms, microscans 2, automatic gain control (AGC) 3 x 10^6 ions, source voltage 4.0 kV, capillary temperature 320 C, and sheath gas 45, auxiliary gas 15, and sweep gas 0 (all nitrogen). Data converted to mzXML using RawConverter. Metabolites were annotated and integrated using Maven in conjunction with the KEGG database."},

"MS_METABOLITE_DATA":{
"Units":"peak area",

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}

}